Hepatic fibrosis or its end-stage form, cirrhosis, is a leading cause of morbidity and mortality both in the United States and worldwide. Alcohol abuse accounts for over 50% of cirrhosis-related deaths. Preliminary data from our laboratory suggests a novel molecular pathway that may contribute to alcoholic cirrhosis, and which could serve as a target for future therapeutic interventions. It has long been known that ethanol causes adenosine release both in vitro and in vivo in humans. In preliminary studies, we have demonstrated that ethanol causes the release of adenosine from hepatocytes and the resulting extracellular adenosine, acting through one of its cell surface receptors (A2A receptor), increases collagen production in rat and human hepatic stellate cells (HSC), the principal fibrogenic cell type in the liver. This adenosine-stimulated collagen production by HSC is blocked by an adenosine A2A receptor antagonist. Furthermore, our results indicate that adenosine-stimulated collagen production is mediated in part by the cAMP-protein kinase A pathway. We now have compelling evidence that adenosine and its receptors mediate hepatic fibrous tissue formation in vivo. First, adenosine A2A receptor-knockout mice are protected from hepatic fibrosis in two widely accepted animal models (carbon tetrachloride- and thioacetamide-induced cirrhosis) (6, 7); and secondly, mice with high circulating and tissue levels of adenosine (adenosine deaminase partially-deficient mice) develop cirrhosis spontaneously. These exciting findings suggested to us that manipulations of the intracellular pathways involved in the deposition of connective tissue in the liver may lead to the discovery of treatments suitable for the prevention or therapy of hepatic fibrosis caused by alcohol consumption, for which there is presently no cure, and perhaps from other causes, whether microbial or autoimmune. In pursuit of this long-term goal, we will test the following hypotheses: Hypothesis I: Modulation of adenosine signaling regulates collagen accumulation in the liver through effects on both collagen production and collagen breakdown. Specific Aim: We will study the effect of adenosine receptor agonists and antagonists on collagen, metalloproteinase, and metalloproteinase inhibitor production in human HSC cell line, LX- 1, and primary rat and human HSC. Hypothesis II: Activation of the adenosine A2A receptor produces hepatic fibrosis in vivo. Specific Aim: Determine whether adenosine, acting at A2A receptors, plays a role in the development of hepatic fibrosis in in vivo murine models (carbon tetrachloride- and thioacetamide-induced cirrhosis; adenosine deaminase-partially deficient mice that spontaneously develop cirrhosis; adenosine A2A or A3 receptor-deficient mice). Hypothesis III: The adenosine A2A receptor modulates collagen production via the cAMP pathway or the MAPK pathway, or both. Specific Aim: Investigate the intracellular mechanisms involved in signaling an increase in collagen production (cAMP pathway, MAPK pathway, CREB phosphorylation).